An unlabelled peptide expression data file called “pep_edata.csv” was uploaded to pmart. The column that designates unique molecules was marked as “Mass_Tag_ID”. The original scale of the data was raw intensity and was changed to log2. The value to denote missing data was “NA” and the expression data was not already normalized.
An associated biomolecule information file was also uploaded called “pep_emeta.csv” and the protein identifier column was designated as “Protein”.
An associated sample information file was also uploaded called “pep_fdata.csv”. Trimmed sample names were not used. The column in the sample information file which indicates sample names was designated as “SampleID”.
This table summarizes all the user specified main effects or covariates in pmart. The “Selected Column Name” denotes the name of the column in the sample information file assigned as a main effect or covariate.
| Main Effect or Covariate | Selected Column Name |
|---|---|
| First Main Effect | Condition |
| Second Main Effect | None selected |
| First Covariate | None selected |
| Second Covariate | None selected |
The first column in the table below denotes a property of the peptide data, and the “Data” column states that property’s value.
| Data | |
|---|---|
| Class | pepData |
| Unique SampleIDs (f_data) | 10 |
| Unique Mass_Tag_IDs (e_data) | 13022 |
| Unique Proteins (e_meta) | 1736 |
| Missing Observations | 35221 |
| Proportion Missing | 0.27 |
| Samples per group: Infection | 7 |
| Samples per group: Mock | 3 |
In the table below, the first column denotes the sample and the second is the missing number of observations. The third column represents the second as a percentage of the total number of observations for that sample.
| Missing Observations | Proportion Missing | |
|---|---|---|
| Infection1 | 2620 | 0.201 |
| Infection3 | 3446 | 0.265 |
| Infection4 | 3978 | 0.305 |
| Infection6 | 3534 | 0.271 |
| Infection7 | 2896 | 0.222 |
| Infection8 | 6015 | 0.462 |
| Infection9 | 4953 | 0.380 |
| Mock1 | 2680 | 0.206 |
| Mock2 | 2550 | 0.196 |
| Mock3 | 2549 | 0.196 |
Filters were applied.
| Filter | Type | Parameters |
|---|---|---|
| Molecule Filter | Biomolecule | Molecule Number Min: 2 |
| CV Filter | Biomolecule | Max CV: 150 |
| imd-ANOVA Filter | Biomolecule | ANOVA Min: 2 & G-test Min: 3 |
| Proteomics Filter | Biomolecule | Peptide Number Min: 2 & Remove Degenerates: Yes |
| rMD Filter | Sample | P-Value Threshold: 0.001 & Outlier Metrics: MAD, Kurtosis, Skewness, Correlation |
| Custom Filter | Sample | The custom filter removedthe following sample(s): Infection5 |
A molecule filter was applied to the data, removing Mass_Tag_IDs not having at least 2 samples.
A coefficient of variation (CV) filter was applied to the data removing Mass_Tag_IDs with a CV greater than 150
An ANOVA filter was applied to the data, removing Mass_Tag_IDs not having at least 2 non-missing values per group. Additionaly, an IMD (independence of missing data) filter was applied to the data, removing Mass_Tag_IDs not having at least 3 non-missing values in at least one of the groups.
A degenerate peptide filter was applied to the data, identifying Mass_Tag_IDs mapping to more than one protein. Additionally, a protein filter was applied to the data, identifying proteins not having at least 2 Mass_Tag_IDs mapping to them.
A robust Mahalanobis distance (rMD) filter was applied to the data, removing SampleIDs with an associated rMD-associated p-value less than 0.001.
The custom filter removedthe following sample(s): Infection5
Peptide data was normalized.
SPANS was run to determine optimum normalization parameters.
Manual normalization was used to normalize the data.
| Attribute | Value |
|---|---|
| Subset Function | los |
| Subset Parameters | los: 0.05 |
| Normalization Function | mean |
The analysis step was run.
The following groups were compared: Infection_vs_Mock. Reported below are the parameters used in the statistical analysis, followed by the number of siginificant biomolecules for each comparison.
| Attribute | Value |
|---|---|
| Test Method | anova |
| Multiple Comparison Adjustment | holm |
| Significance Threshold | 0.05 |
| Comparison | Up_total | Down_total | Up_anova | Down_anova | Up_gtest | Down_gtest | |
|---|---|---|---|---|---|---|---|
| Infection_vs_Mock | Infection_vs_Mock | 1509 | 2742 | 1509 | 2742 | 0 | 0 |